Hydrophilic monolith based immobilized enzyme reactors in capillary and on microchip for high-throughput proteomic analysis

Liang, Yu; Tao, Dingyin; Ma, Junfeng; Sun, Liangliang; Liang, Zhen; Zhang, Lihua; Zhang, Yukui

doi:10.1016/j.chroma.2011.02.073

Cited by 50 publications

(34 citation statements)

References 30 publications

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“…Keunggulan penggunaan metode ini dapat meningkatkan stabilitas enzim, meningkatkan volume spesifik enzim yang terikat pada material pendukung (enzyme loading), dan bersifat reusable atau dapat digunakan kembali (Chen et al, 2011;Shangguan et al, 2015). Imobilisasi enzim pada suatu mikroreaktor juga memiliki efisiensi pemecahan (digestion) tinggi dan dapat dengan mudah dihubungkan dengan sistem pemisahan dan deteksi secara otomatis (Liang et al, 2011).…”

Section: Pendahuluanunclassified

“…Studi mengenai proteomik berfokus pada karakterisasi biomolekul yang berbeda. Selain itu, studi ini juga berkaitan dengan proses identifikasi keseluruhan protein dalam sel, dan jaringan (Liang et al, 2011). Pada awal pengembangannya, studi mengenai protein digestion dilakukan menggunakan metode konvensional (offline).…”

Section: Pendahuluanunclassified

“…Pada dasarnya, metode pemisahan dalam larutan masih belum efisien terutama dalam aplikasinya untuk penggunaan rutin. Seiring dengan berkembangnya ilmu pengetahuan metode tersebut mulai ditinggalkan karena dinilai memiliki kelemahan yaitu waktu digestion lama, autodigestion pada enzim tidak dapat dihindari, reproduksibilitas rendah dan berlangsung secara offline sehingga distribusi enzim dalam larutan tidak merata (Liang et al, 2011;Safdar et al, 2014). Hal tersebut sangat berbanding terbalik dengan metode online digestion, dimana pada metode tersebut semua prosesnya berlangsung secara online menggunakan sistem HPLC dalam sebuah kolom mikroreaktor sehingga efisiensi waktu pemisahan meningkat karena hanya berlangsung dalam hitungan menit.…”

Section: Pendahuluanunclassified

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Preparasi dan Karakterisasi Nanobiokatalis Mikroreaktor dengan Polimer Monolitik Nanopori Enzim Termobilisasi

Anggraeny

Raeni

Iftitah

et al. 2018

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Nanobiocatalyst microreactor column of an immobilized trypsin onto nanoporous monolithic polymer has been developed. In this work, a poly (glycidyl methacrylate-co-ethylene glycol dimethacrylate) (poly(GMA-co-EDMA)) monolih with total monomer %T 40 and cross-linker %C 25 was prepared as a nanoporous polymeric-support by in-situ copolymerization inside the microbore silicosteel tubing (1.0 mm i.d x 100 mm). This nanoporous monolithic-support was chemically modified with trypsin through the ring-opening reaction of epoxide group to form a nanobiocatalyst microreactor. In order to obtain reliable microreactor, some parameters such as immobilization time (amount of trypsin), and effect of glutaraldehyde activation on the trypsin-immobilized monolithic column were optimized. The pore size distribution, morphology and elemental composition of the monolith were studied by inverse size exclusion chromatography (ISEC) and scanning electron microscopy (SEM-EDX), respectively. While permeability of the monolith was obtained through measurement of its back pressure using HPLC pump. The monolith was predominated by mesoporous content (65.85%) while the flow-through pore was 28.20%. From SEM-EDX, it was found that the highest nitrogen content (12.40%), which indicates the amount of trypsin chemically attached on the monolitic support, was found in the nanobiocatalyst microreactor with immobilization time 4h without glutaraldehyde as activation agent. The optimized nanobiocatalyst microreactor column was successfully applied to a rapid and efficient digestion of protein samples in the range of second to minute.Keywords: nanoporous monolit, immobilized enzyme, nanobiocatalyst microreactor, protein digestion Pada penelitian ini telah dilakukan pengembangan pembuatan kolom nanobiokatalis mikroreaktor yang terimobilisasi oleh enzim tripsin pada polimer monolitik nanopori. Polimer monolitik nanopori dibuat secara insitu kopolimerisasi dalam kolom silicosteel (1.0 mm i.d x 100 mm) dari monomer GMA dan EDMA sebagai crosslinker serta penambahan porogen dengan komposisi 1-propanol/1,4-butanediol/air (7:4:1) dan inisiator radikal AIBN 1%. Proses polimerisasi berlangsung selama 12 jam pada suhu 60°C dengan persentase total monomer %T 40 dan persentase crosslinker %C 25. Monolit dimodifikasi secara kimia menggunakan metode imobilisasi enzim melalui reaksi pembukaan cincin *Penulis korespondensi

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Section: Pendahuluanunclassified

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Preparasi dan Karakterisasi Nanobiokatalis Mikroreaktor dengan Polimer Monolitik Nanopori Enzim Termobilisasi

Anggraeny

Raeni

Iftitah

et al. 2018

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“…Liang et al 67 formed a hydrophilic monolith on a microchip by photopolymerization of Nacryloxysuccinimide and PEGDA for immobilization of trypsin. This reactor was coupled with MS through a C18 particle-packed electrospray emitter tip ( Figure 5(a)); BSA, myoglobin, and cytochrome C were digested, separated, and identified using this setup with 50%-60% sequence coverage.…”

Section: Proteolytic Enzyme Reactorsmentioning

confidence: 99%

Advances in monoliths and related porous materials for microfluidics

2016

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In recent years, the use of monolithic porous polymers has seen significant growth. These materials present a highly useful support for various analytical and biochemical applications. Since their introduction, various approaches have been introduced to produce monoliths in a broad range of materials. Simple preparation has enabled their easy implementation in microchannels, extending the range of applications where microfluidics can be successfully utilized. This review summarizes progress regarding monoliths and related porous materials in the field of microfluidics between 2010 and 2015. Recent developments in monolith preparation, solid-phase extraction, separations, and catalysis are critically discussed. Finally, a brief overview of the use of these porous materials for analysis of subcellular and larger structures is given.

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“…More importantly, some IMERs could be readily coupled to separation and identification systems, enabling fast, efficient, high-throughput and automated proteome analysis [7]. Till now, enzymes have been immobilized on different supports, such as membranes [8], carbon materials [9], monolithic materials [10][11][12][13][14][15][16][17], polymers [18][19][20][21], silica materials [22,23] and hybrids [24]. However, the improved binding capacity of enzymes and the decreased peptide residue on matrices are still the hot-points under serious consideration in this field.…”

Section: Introductionmentioning

confidence: 99%

Hydrophilic immobilized trypsin reactor with magnetic graphene oxide as support for high efficient proteome digestion

Jiang

Yang

Zhao

et al. 2012

Journal of Chromatography A

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Hydrophilic monolith based immobilized enzyme reactors in capillary and on microchip for high-throughput proteomic analysis

Cited by 50 publications

References 30 publications

Preparasi dan Karakterisasi Nanobiokatalis Mikroreaktor dengan Polimer Monolitik Nanopori Enzim Termobilisasi

Preparasi dan Karakterisasi Nanobiokatalis Mikroreaktor dengan Polimer Monolitik Nanopori Enzim Termobilisasi

Advances in monoliths and related porous materials for microfluidics

Hydrophilic immobilized trypsin reactor with magnetic graphene oxide as support for high efficient proteome digestion

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